Benzo (k) fluoranthene (BkF) has adverse effects on male reproduction, but its specific mechanism of action is still unclear. This study focused on the role of RNA reading protein YTHDF2 and its mechanism in BkF induced male reproductive injury. Mouse GC-2 spermatocytes were exposed to 0, 40, 80, 160 μM BkF. It was found that BkF significantly increased the apoptosis of GC-2 cell and decreased its survival rate. BCL2 in spermatocytes decreased significantly, while the expression of P53 and BAX exhibited a notable increase. Interestingly, the expression of RNA reading protein YTHDF2 progressively rose in tandem with the escalating BkF exposure dosage. Overexpression of YTHDF2 significantly reduced the viability of cells and increased the apoptosis rate. Meanwhile, there was a substantial increase in the expression of P53 and BAX, BCL2 was significantly down-regulated. On the contrary, interfering with YTHDF2 increased cell proliferation and reduced cell apoptosis. Furthermore, YTHDF2 overexpression exacerbated the decrease in cell viability under BkF exposure, while YTHDF2 knockdown was the opposite. The results from the RIP assay demonstrated a significant enhancement in the interaction of YTHDF2 protein with BCL2 mRNA following the overexpression of YTHDF2. In addition, animal experiments showed that there was an increase in apoptosis and a decrease in proliferation of testicular cells in mice in the high-dose (30 mg/kg) BkF group by TUNEL staining and Ki67 staining. Immunohistochemical analysis showed that BCL2 levels were significantly lower in the high-dose group than in the control group, while YTHDF2, P53 and BAX were dramatically increased. In summary, our study suggests that YTHDF2 has been implicated in BkF-induced male reproductive injury by promoting the degradation of BCL2.

中文翻译：

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RNA阅读蛋白YTHDF2通过调节BCL2的稳定性介导苯并荧蒽诱导的男性生殖损伤


苯并(k)荧蒽(BkF)对男性生殖有不良影响，但其具体作用机制尚不清楚。本研究重点探讨RNA阅读蛋白YTHDF2在BkF诱导的男性生殖损伤中的作用及其机制。将小鼠 GC-2 精母细胞暴露于 0、40、80、160 μM BkF。结果发现BkF显着增加GC-2细胞的凋亡并降低其存活率。精母细胞中BCL2的表达显着下降，而P53和BAX的表达显着增加。有趣的是，随着 BkF 暴露剂量的增加，RNA 阅读蛋白 YTHDF2 的表达逐渐上升。 YTHDF2的过表达显着降低了细胞的活力并增加了细胞凋亡率。同时，P53和BAX的表达大幅增加，BCL2的表达显着下调。相反，干扰 YTHDF2 会增加细胞增殖并减少细胞凋亡。此外，YTHDF2 过表达加剧了 BkF 暴露下细胞活力的下降，而 YTHDF2 敲低则相反。 RIP 测定的结果表明，在 YTHDF2 过表达后，YTHDF2 蛋白与 BCL2 mRNA 的相互作用显着增强。此外，动物实验通过TUNEL染色和Ki67染色显示，高剂量（30 mg/kg）BkF组小鼠睾丸细胞凋亡增加，增殖减少。免疫组织化学分析显示，高剂量组的BCL2水平显着低于对照组，而YTHDF2、P53和BAX则显着升高。 总之，我们的研究表明，YTHDF2 通过促进 BCL2 的降解而参与 BkF 诱导的男性生殖损伤。